This invention relates to a novel class of cyclic diaza compounds which are potent and highly selective inhibitors of factor Xa or factor Xa when assembled in the prothrombinase complex. These compounds show selectivity for factor Xa versus other proteases of the coagulation (e.g. thrombin, fVIIa, fIXa) or the fibrinolytic cascades (e.g. plasminogen activators, plasmin).
Blood coagulation protects mammalian species when the integrity of the blood vessel wall is damaged and uncontrolled loss of blood threatens survival. Coagulation, resulting in the clotting of blood, is an important component of hemostasis. Under normal hemostatic circumstances, there is maintained an acute balance of clot formation and clot, removal (fibrinolysis). The blood coagulation cascade involves the conversion of a variety of inactive enzymes (zymogens) into active enzymes, which ultimately convert the soluble plasma protein fibrinogen into an insoluble matrix of highly cross-linked fibrin. (See Davie, et al., xe2x80x9cThe Coagulation Cascade: Initiation, Maintenance and Regulationxe2x80x9d Biochemistry 30:10363-10370 (1991)). Blood platelets which adhere to damaged blood vessels are activated and incorporated into the clot and thus play a major role in the initial formation and stabilization of hemostatic xe2x80x9cplugsxe2x80x9d. In certain diseases of the cardiovascular system, deviations from normal hemostasis push the balance of clot formation and clot dissolution towards life-threatening thrombus formation when thrombi occlude blood flow in coronary vessels (myocardial infarctions) or limb and pulmonary veins (venous thrombosis). Although platelets and blood coagulation are both involved in thrombus formation, certain components of the coagulation cascade are primarily responsible for the amplification or acceleration of the processes involved in platelet aggregation and fibrin deposition.
A key enzyme in the coagulation cascade, as well as in hemostasis, is thrombin. Thrombin is intimately involved in the process of thrombus formation, but under normal circumstances can also play an anticoagulant role in hemostasis through its ability to convert protein C into activated protein C in a thrombomodulin-dependent manner. Thrombin plays a central role in thrombosis through its ability to catalyze the penultimate conversion of fibrinogen into fibrin and through its potent platelet activation activity. Direct or indirect inhibition of thrombin activity has been the focus of a variety of recent anticoagulant strategies as reviewed by Claeson xe2x80x9cSynthetic Peptides and Peptidomimetics as Substrates and Inhibitors of Thrombin and Other Proteases in the Blood Coagulation Systemxe2x80x9d, Blood Coag. Fibrinol. 5:411-436 (1994). The major classes of anticoagulants currently used in the clinic directly or indirectly affect thrombin (i.e. heparins, low-molecular weight heparins and coumarins). Thrombin is generated at the convergence of the intrinsic and extrinsic coagulation pathways by the prothrombinase complex. The prothrombinase complex is formed when activated Factor X (factor Xa) and its non-enzymatic cofactor, factor Va assemble on phospholipid surfaces in a Ca+2-dependent fashion as reviewed by Mann, et al., xe2x80x9cSurface-Dependent Reactions of the Vitamin K-Dependent Enzymesxe2x80x9d, Blood 76:1-16 (1990). The prothrombinase complex converts the zymogen prothrombin into the active procoagulant thrombin.
The location of the prothrombinase complex at the convergence of the intrinsic and extrinsic coagulation pathways, and the significant amplification of thrombin generation (393,000-fold over uncomplexed factor Xa) mediated by the complex at a limited number of targeted catalytic units present at vascular lesion sites, suggests that inhibition of thrombin generation is an ideal method to block uncontrolled procoagulant activity. Unlike thrombin, which acts on a variety of protein substrates as well as at a specific receptor, factor Xa appears to have a single physiologic substrate, namely prothrombin.
Plasma contains an endogenous inhibitor of both the factor VIIa-tissue factor (TF) complex and factor Xa called tissue factor pathway inhibitor (TFPI). TFPI is a Kunitz-type protease inhibitor with three tandem Kunitz domains. TFPI inhibits the TF/fVIIa complex in a two-step mechanism which includes the initial interaction of the second Kunitz domain of TFPI with the active site of factor Xa, thereby inhibiting the proteolytic activity of factor Xa. The second step involves the inhibition of the TF/fVIIa complex by formation of a quaternary complex TF/fVIIa/TFPI/fXa as described by Girard, et al., xe2x80x9cFunctional Significance of the Kunitz-type Inhibitory Domains of Lipoprotein-associated Coagulation Inhibitorxe2x80x9d, Nature 338:518-520 (1989).
Polypeptides derived from hematophagous organisms have been reported which are highly potent and specific inhibitors of factor Xa. U.S. Pat. No. 4,588,587 awarded to Gasic, describes anticoagulant activity in the saliva of the Mexican leech, Haementeria officinalis. A principal component of this saliva is shown to be the polypeptide factor Xa inhibitor, antistasin, by Nutt, et al., xe2x80x9cThe Amino Acid Sequence of Antistasin, a Potent Inhibitor of Factor Xa Reveals a Repeated Internal Structurexe2x80x9d, J. Biol. Chem. 263:10162-10167 (1988).
Another potent and highly specific inhibitor of Factor Xa, tick anticoagulant peptide, has been isolated from the whole body extract of the soft tick Ornithidoros moubata, as reported by Waxman, et al., xe2x80x9cTick Anticoagulant Peptide (TAP) is a Novel Inhibitor of Blood Coagulation Factor Xaxe2x80x9d, Science 248:593-596 (1990).
Other polypeptide type inhibitors of factor Xa have been reported including the following citations by: Condra, et al., xe2x80x9cIsolation and Structural Characterization of a Potent Inhibitor of Coagulation Factor Xa from the Leech Haementeria ghilianiixe2x80x9d, Thromb. Haemost. 61:437-441 (1989); Blankenship, et al., xe2x80x9cAmino Acid Sequence of Ghilanten: Anti-coagulant-antimetastatic Principle of the South American Leech, Haementeria ghilianiixe2x80x9d, Biochem. Biophys. Res. Commun. 166:1384-1389 (1990); Brankamp, et al., xe2x80x9cGhilantens: Anticoagulants, Antimetastatic Proteins from the South American Leech Haementeria ghilianiixe2x80x9d, J. Lab. Clin. Med. 115:89-97 (1990); Jacobs, et al., xe2x80x9cIsolation and Characterization of a Coagulation Factor Xa Inhibitor from Black Fly Salivary Glandsxe2x80x9d, Thromb. Haemost. 64:235-238 (1990); Rigbi, et al., xe2x80x9cBovine Factor Xa Inhibiting Factor and Pharmaceutical Compositions Containing the Samexe2x80x9d, European Patent Application, 352,903 (1990); Cox, xe2x80x9cCoagulation Factor X Inhibitor From the Hundred-pace Snake Deinagkistrodon acutus venomxe2x80x9d, Toxicon 31: 1445-1457 (1993); Cappello, et al., xe2x80x9cAncylostoma Factor Xa Inhibitor: Partial Purification and its Identification as a Major Hookworm-derived Anticoagulant In Vitroxe2x80x9d, J. Infect. Dis. 167:1474-1477 (1993); Seymour, et al., xe2x80x9cEcotin is a Potent Anticoagulant and Reversible Tight-binding Inhibitor of Factor Xaxe2x80x9d, Biochemistry 33:3949-3958 (1994).
Factor Xa inhibitory compounds which are not large polypeptide-type inhibitors have also been reported including: Tidwell, et al., xe2x80x9cStrategies for Anticoagulation With Synthetic Protease Inhibitors. Xa Inhibitors Versus Thrombin Inhibitorsxe2x80x9d, Thromb. Res. 19:339-349 (1980); Turner, et al., xe2x80x9cp-Amidino Esters as Irreversible Inhibitors of Factor IXa and Xa and Thrombinxe2x80x9d, Biochemistry 25:4929-4935 (1986); Hitomi, et al., xe2x80x9cInhibitory Effect of New Synthetic Protease Inhibitor (FUT-175) on the Coagulation Systemxe2x80x9d, Haemostasis 15:164-168 (1985); Sturzebecher, et al., xe2x80x9cSynthetic Inhibitors of Bovine Factor Xa and Thrombin. Comparison of Their Anticoagulant Efficiencyxe2x80x9d, Thromb. Res. 54:245-252 (1989); Kam, et al., xe2x80x9cMechanism Based Isocoumarin Inhibitors for Trypsin and Blood Coagulation Serine Proteases: New Anticoagulantsxe2x80x9d, Biochemistry 27:2547-2557 (1988); Hauptmann, et al., xe2x80x9cComparison of the Anticoagulant and Antithrombotic Effects of Synthetic Thrombin and Factor Xa Inhibitorsxe2x80x9d, Thromb. Haemost. 63:220-223 (1990); Miyadera, et al., Japanese Patent Application JP 6327488 (1994); Nagahara, et al., xe2x80x9cDibasic (Amidinoaryl)propanoic Acid Derivatives as Novel Blood Coagulation Factor Xa Inhibitorsxe2x80x9d, J. Med. Chem. 37:1200-1207 (1994); Vlasuk, et al., xe2x80x9cInhibitors of Thrombosisxe2x80x9d, WO 93/15756; and Brunck, et al., xe2x80x9cNovel Inhibitors of Factor Xaxe2x80x9d, WO 94/13693. Al-obeidi, et al., xe2x80x9cFactor Xa Inhibitorsxe2x80x9d, WO 95/29189, discloses pentapeptide X1-Y-I-R-X2 derivatives as factor Xa inhibitors. Said compounds are useful for inhibiting blood clotting in the treatment of thrombosis, stroke, and myocardial infarction.
The present invention relates to novel peptide mimetic analogs, their pharmaceutically acceptable isomers, salts, hydrates, solvates and prodrug derivatives.
In another aspect, the present invention includes pharmaceutical compositions comprising a pharmaceutically effective amount of the compounds of this invention and a pharmaceutically acceptable carrier. These compositions are useful as potent and specific inhibitors of blood coagulation in mammals.
In yet another aspect, the invention relates to methods of using these inhibitors as therapeutic agents for disease states in mammals which have disorders of coagulation such as in the treatment or prevention of unstable angina, refractory angina, myocardial infarction, transient ischemic attacks, thrombotic stroke, embolic stroke, disseminated intravascular coagulation including the treatment of septic shock, deep venous thrombosis in the prevention of pulmonary embolism or the treatment of reocclusion or restenosis of reperfused coronary arteries. These compositions may optionally include anticoagulants, antiplatelet agents, and thrombolytic agents.
In other aspects of the invention compounds are provided which are useful as diagnostic reagents.
In preferred embodiments, the present invention provides compounds of general formula I: 
Wherein:
R1 is selected from the group consisting of H, C1-6alkyl, C3-8cycloalkyl, C1-3alkylaryl, C1-3alkyl-C3-8cycloalkyl and aryl and R2 is H, or R1 and R2 are taken together to form a carbocyclic ring;
m is an integer from 0-2;
n is an integer from 0-6;
k is an integer from 0-1;
p is an integer from 0-1;
q is an integer from 0-3;
r is an integer from 0-4;
s is an integer from 0-1;
t is an integer from 0-4;
A is selected from the group consisting of R3, xe2x80x94NR3R4, 
where
R3, R4, R5 and R6 are independently selected from the group consisting of H, xe2x80x94OH, C1-6alkyl, aryl and C1-4alkylaryl; R7 is selected from the group consisting of H, C1-6alkyl, aryl and C1-4alkylaryl, or can be taken together with R5 or R6 to form a 5-6 membered ring; and R8 is selected from the group consisting of H, C1-6alkyl, aryl and C1-4alkylaryl, or can be taken together with R6 to form a 5-6 membered ring;
Q is selected from the group consisting of a direct link, xe2x80x94Sxe2x80x94, xe2x80x94SOxe2x80x94, xe2x80x94SO2xe2x80x94, xe2x80x94Oxe2x80x94, xe2x80x94SO2NHxe2x80x94, xe2x80x94NHSO2xe2x80x94, xe2x80x94COOxe2x80x94, xe2x80x94CONHxe2x80x94, C1-6alkyl, C3-8cycloalkyl, C2-6alkenyl, C2-6alkenylaryl, aryl and a five to ten membered heterocyclic ring system containing 1-4 heteroatoms selected from the group consisting of N, O and S;
D is selected from the group consisting of a direct link, xe2x80x94COxe2x80x94, xe2x80x94SO2xe2x80x94, xe2x80x94Oxe2x80x94COxe2x80x94, xe2x80x94NR9xe2x80x94SO2xe2x80x94 or xe2x80x94NR9xe2x80x94COxe2x80x94, where R9 is H, xe2x80x94OH, C1-6alkyl, aryl or C1-4alkylaryl;
E is selected from the group consisting of a direct link, C3-8cycloalkyl, aryl, or a five to ten membered heterocyclic ring system containing 1-4 heteroatoms selected from the group consisting of N, O and S;
G is selected from the group consisting of R10, xe2x80x94NR10OR11, 
where
R10, R11, R12 and R13 are independently selected from the group consisting of H, xe2x80x94OH, C1-6alkyl, aryl and C1-4alkylaryl; R14 is selected from the group consisting of H, C1-6alkyl, aryl and C1-4alkylaryl, or can be taken together with R12 or R13 to form a 5-6 membered ring; and R15 is selected from the group consisting of H, C1-6alkynyl, aryl and C1-4alkylaryl, or can be taken together with R13 to form a 5-6 membered ring; with the proviso that when G is R10, then E must contain at least one N atom;
K is selected from the group consisting of a direct link, C3-8cycloalkyl, C2-6alkenyl, C2-6alkenylaryl, aryl and a five to ten membered heterocyclic ring system containing 1-4 heteroatoms selected from the group consisting of N, O and S;
T is selected from the group consisting of R28, xe2x80x94NR28R29, 
where
R28, R29, R30 and R31 are independently selected from the group consisting of H, xe2x80x94OH, C1-6alkyl, aryl and C1-4alkylaryl; R32 is selected from the group consisting of H, C1-6alkyl, aryl and C1-4alkylaryl, or can be taken together with R30 or R31 to form a 5-6 membered ring; and R33 is selected from the group consisting of H, C1-6alkyl, aryl and C1-4alkylaryl, or can be taken together with R31 to form a 5-6 membered ring;
X and Y are independently selected from the group consisting of O and H2;
W is selected from the group consisting of H, 
where
R16 and R17 are independently selected from the group consisting of H, C1-3alkyl and aryl; and Z is selected from the group consisting of H, xe2x80x94COOR18, xe2x80x94CONR18R19, xe2x80x94CF3, xe2x80x94CF2CF3 and a group having the formula: 
where:
R18 and R19 are independently selected from the group consisting of H, C1-6alkyl, aryl and C1-4alkylaryl;
U is selected from the group consisting of xe2x80x94Oxe2x80x94, xe2x80x94Sxe2x80x94, xe2x80x94Nxe2x80x94 and xe2x80x94NHxe2x80x94; and
V is selected from the group consisting of xe2x80x94Oxe2x80x94, xe2x80x94Sxe2x80x94, xe2x80x94Nxe2x80x94 and xe2x80x94NHxe2x80x94; with the proviso that at least one of U or V is xe2x80x94Nxe2x80x94 or xe2x80x94NHxe2x80x94;
R20 is selected from the group consisting of H, C1-6alkyl, C2-6alkenyl, C0-6alkylaryl, C2-6alkenylaryl, C0-6alkylheterocyclo, C2-6alkenylheterocyclo, xe2x80x94CF3 and xe2x80x94CF2CF3;
J is selected from the group consisting of xe2x80x94Sxe2x80x94, xe2x80x94SOxe2x80x94, xe2x80x94SO2xe2x80x94, xe2x80x94Oxe2x80x94 and xe2x80x94NR21xe2x80x94, where R21is selected from the group consisting of H, C1-6alkyl and benzyl; and
L is selected from the group consisting of: 
xe2x80x83a C6-10 heterocyclic ring system substituted by R24 and R25 and containing 1-4 heteroatoms selected from N, S and O;
where d is an integer from 0-2;
R22 and R23 are independently selected from the group consisting of H, C1-6alkyl, aryl, C1-6alkylaryl, xe2x80x94COOR26, xe2x80x94CONR26R27, xe2x80x94CN and xe2x80x94CF3;
R24 and R25 are independently selected from the group consisting of H, C1-6alkyl, aryl, C1-6alkylaryl, C1-4alkyloxy, halogen, xe2x80x94NO2, xe2x80x94NR26R27, xe2x80x94NR26COR27, xe2x80x94OR26, xe2x80x94OCOR26, xe2x80x94COOR26, xe2x80x94CONR26R27, xe2x80x94CN, xe2x80x94CF3, xe2x80x94SO2NR26R27 and C1-6alkyl-OR26; and
R26 and R27 are independently selected from the group consisting of H, C1-6alkyl, C1-3alkylaryl and aryl; and all pharmaceutically acceptable salts and optical isomers thereof.
In accordance with the present invention and as used herein, the following terms are defined with the following meanings, unless explicitly stated otherwise.
The term xe2x80x9calkylxe2x80x9d refers to saturated aliphatic groups including straight-chain, branched-chain, cyclic groups, and combinations thereof, having the number of carbon atoms specified, or if no number is specified, having up to 12 carbon atoms. The term xe2x80x9ccycloalkylxe2x80x9d refers to a mono-, bi-, or tricyclic aliphatic ring having 3 to 12 carbon atoms, preferably 3 to 7 carbon atoms.
The term xe2x80x9calkenylxe2x80x9d refers to unsaturated aliphatic groups including straight-chain, branched-chain, cyclic groups, and combinations thereof, having at least one double bond and having the number of carbon atoms specified.
The term xe2x80x9carylxe2x80x9d refers to an unsubstituted or substituted aromatic ring(s), substituted with one, two or three substituents such as, by way of example and not limitation, C1-6alkoxy, C1-6 alkyl, C1-6 alkylamino, hydroxy, halogen, cyano (xe2x80x94CN), mercapto, nitro (xe2x80x94NO2), thioalkoxy, carboxaldehyde, carboxyl, carboalkoxy, carboxamide, xe2x80x94NRxe2x80x2Rxe2x80x3, xe2x80x94NRxe2x80x2CORxe2x80x3, xe2x80x94OR, xe2x80x94OCOR, xe2x80x94COOR, xe2x80x94CONRxe2x80x2Rxe2x80x3, xe2x80x94CF3, xe2x80x94SO2NRxe2x80x2Rxe2x80x3 and C1-6alkyl-OR; aryl, C1-6alkylaryl (where the R groups can be H, C1-6alkyl, C1-3alkylaryl and aryl), including but not limited to carbocyclic aryl, heterocyclic aryl, biaryl and triaryl groups and the like, all of which may be optionally substituted. Preferred aryl groups include phenyl, halophenyl, C1-6 alkylphenyl, naphthyl, biphenyl, phenanthrenyl, naphthacenyl, and aromatic heterocyclics or heteroaryls, the latter of which is an aryl group containing one to four heteroatoms selected from the group consisting of nitrogen, oxygen and sulfur. Aryl groups preferably have 5-14 carbon atoms making up the ring(s) structure, while heteroaryls preferably have 1-4 heteroatoms, with the remaining 4-10 atoms being carbon atoms.
The terms xe2x80x9cheterocycloxe2x80x9d and xe2x80x9chetero cyclic ring systemxe2x80x9d as used herein refer to any saturated or unsaturated mono- or bicyclic ring system, containing from one to four heteroatoms, selected from the group consisting of nitrogen, oxygen and sulfur. Typical examples of monocyclic ring systems include piperidinyl, pyrrolidinyl, pyridinyl, piperidonyl, pyrrolidonyl and thiazolyl, while examples of bicyclic ring systems include benzimidazolyl, benzothiazolyl and benzoxazolyl, all of which may be substituted.
The term xe2x80x9ccarbocyclic ringxe2x80x9d as used herein refers to any saturated or unsaturated ring containing from three to six carbon atoms.
The terms xe2x80x9calkylarylxe2x80x9d and xe2x80x9calkenylarylxe2x80x9d as used herein refer to an alkyl group or alkenyl group, respectively, having the number of carbon atoms designated, appended to one, two, or three aryl groups. The term benzyl as used herein refers to xe2x80x94CH2xe2x80x94C6H5 (C1alkylaryl).
The term xe2x80x9calkyloxyxe2x80x9d as used herein refers to an alkyl group linked to an oxygen atom, such as methoxy, ethoxy, and so forth.
The term xe2x80x9chalogenxe2x80x9d as used herein refer to Cl, Br, F or I substituents.
The term xe2x80x9cdirect linkxe2x80x9d as used herein refers to a bond directly linking the substituents on each side of the direct link. When two adjacent substituents are defined as each being a xe2x80x9cdirect linkxe2x80x9d, it is considered to be a single bond.
Two substituents are xe2x80x9ctaken together to form a 5-6 membered ringxe2x80x9d means that an ethylene or a propylene bridge, respectively, is formed between the two substituents.
The term xe2x80x9cpharmaceutically acceptable saltsxe2x80x9d includes salts of compounds derived from the combination of a compound and an organic or inorganic acid. These compounds are useful in both free base and salt form. In practice, the use of the salt form amounts to use of the base form; both acid and base addition salts are within the scope of the present invention.
xe2x80x9cPharmaceutically acceptable acid addition saltxe2x80x9d refers to those salts which retain the biological effectiveness and properties of the free bases and which are not biologically or otherwise undesirable, formed with inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid and the like, and organic acids such as acetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, salicylic acid and the like.
xe2x80x9cPharmaceutically acceptable base addition saltsxe2x80x9d include those derived from inorganic bases such as sodium, potassium, lithium, ammonium, calcium, magnesium, iron, zinc, copper, manganese, and aluminum bases, and the like. Particularly preferred are the ammonium, potassium, sodium, calcium and magnesium salts. Salts derived from pharmaceutically acceptable organic nontoxic bases include salts of primary, secondary, and tertiary amines, substituted amines including naturally occurring substituted amines, cyclic amines and basic ion exchange resins, such as isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, ethanolamine, 2-diethylaminoethanol, trimethamine, dicyclohexylamine, lysine, arginine, histidine, caffeine, procaine, hydrabamine, choline, betaine, ethylenediamine, glucosamine, methylglucamine, theobromine, purines, piperizine, piperidine, N-ethylpiperidine, polyamine resins and the like. Particularly preferred organic nontoxic bases are isopropylamine, diethylamine, ethanolamine, trimethamine, dicyclohexylamine, choline, and caffeine.
xe2x80x9cBiological propertyxe2x80x9d for the purposes herein means an in vivo effector or antigenic function or activity that is directly or indirectly performed by a compound of this invention. Effector functions include receptor or ligand binding, any enzyme activity or enzyme modulatory activity, any carrier binding activity, any hormonal activity, any activity in promoting or inhibiting adhesion of cells to an extracellular matrix or cell surface molecules, or any structural role. Antigenic functions include possession of an epitope or antigenic site that is capable of reacting with antibodies raised against it. The biological properties of the compounds of the present invention can be readily characterized by the methods described in Examples 44 and 45 and by such other methods as are well known in the art.
In addition, the following abbreviations are used in this application:
xe2x80x9cBnxe2x80x9d refers to benzyl.
xe2x80x9cBocxe2x80x9d refers to t-butoxycarbonyl.
xe2x80x9cBOPxe2x80x9d refers to benzotriazol-1-yloxy-tris-(dimethylamino) phosphonium hexafluorophosphate.
xe2x80x9cBuxe2x80x9d refers to butyl.
xe2x80x9cCbzxe2x80x9d refers to carbobenzoxy.
xe2x80x9cDCMxe2x80x9d refers to dichloromethane.
xe2x80x9cDIEAxe2x80x9d refers to diisopropylethylamine.
xe2x80x9cDMFxe2x80x9d refers to N,N-dimethylformamide.
xe2x80x9cEtxe2x80x9d refers to ethyl.
xe2x80x9cEt2Oxe2x80x9d refers to diethyl ether.
xe2x80x9cEtOAcxe2x80x9d refers to ethyl acetate.
xe2x80x9cHFxe2x80x9d refers to hydrogen fluoride.
xe2x80x9cMexe2x80x9d refers to methyl.
xe2x80x9cPhxe2x80x9d refers to phenyl.
xe2x80x9cTFAxe2x80x9d refers to trifluoroacetic acid.
xe2x80x9cTHFxe2x80x9d refers to tetrahydrofuran.
xe2x80x9cTosxe2x80x9d refers to p-toluenesulfonyl.
In the compounds of this invention, carbon atoms bonded to four non-identical substituents are asymmetric. Accordingly, the compounds may exist as diastereoisomers, enantiomers or mixtures thereof. The syntheses described herein may employ racemates, enantiomers or diastereomers as starting materials or intermediates. Diastereomeric products resulting from such syntheses may be separated by chromatographic or crystallization methods, or by other methods known in the art. Likewise, enantiomeric product mixtures may be separated using the same techniques or by other methods known in the art. Each of the asymmetric carbon atoms, when present in the compounds of this invention, may be in one of two configurations (R or S) and both are within the scope of the present invention. In the processes described above, the final products may, in some cases, contain a small amount of diastereomeric or enantiomeric products; however, these products do not affect their therapeutic or diagnostic application.
In all of the peptides of the invention, one or more amide linkages (xe2x80x94COxe2x80x94NHxe2x80x94) may optionally be replaced with another linkage which is an isostere such as xe2x80x94CH2NHxe2x80x94, xe2x80x94CH2Sxe2x80x94, xe2x80x94CH2xe2x80x94Oxe2x80x94, xe2x80x94CH2CH2xe2x80x94, xe2x80x94CHxe2x95x90CHxe2x80x94 (cis and trans), xe2x80x94COCH2xe2x80x94, xe2x80x94CH(OH)CH2xe2x80x94, xe2x80x94CH2SOxe2x80x94, and xe2x80x94CH2SO2xe2x80x94. This replacement can be made by methods known in the art. The following references describe preparation of peptide analogs which include these alternative-linking moieties: Spatola, xe2x80x9cPeptide Backbone Modificationsxe2x80x9d (general review) Vega Data, Vol. 1, Issue 3, (March 1983); Spatola, xe2x80x9cChemistry and Biochemistry of Amino Acids, Peptides and Proteins,xe2x80x9d (general review) B. Weinstein, eds., Marcel Dekker, New York, p. 267 (1983); Morley, Trends Pharm. Sci. (general review) pp. 463-468 (1980); Hudson, et al., Int. J. Pept. Prot. Res. 14:177-185 (1979) (xe2x80x94CH2NHxe2x80x94, xe2x80x94CH2CH2xe2x80x94); Spatola, et al., Life Sci. 38:1243-1249 (1986) (xe2x80x94CH2xe2x80x94S); Hann, J. Chem. Soc. Perkin Trans. I pp.307-314 (1982) (xe2x80x94CHxe2x95x90CHxe2x80x94, cis and trans); Almquist, et al., J. Med. Chem. 23:1392-1398 (1980) (xe2x80x94COCH2xe2x80x94); Jennings-White, et al., Tetrahedron Lett. 23:2533 (xe2x80x94COCH2xe2x80x94) (1982); Szelke, et al., European Application EP 45665; CA:97:39405 (1982) (xe2x80x94CH(OH)CH2xe2x80x94); Holladay, et al., Tetrahedron Lett 24:4401-4404 (1983) (xe2x80x94CH(OH)CH2xe2x80x94); and Hruby, Life Sci. 31:189-199 (1982) (xe2x80x94CH2xe2x80x94Sxe2x80x94).
This invention relates to a new class of cyclic diaza compounds selected from those of general formula I which are potent and specific inhibitors of Xa, their pharmaceutically acceptable compositions thereof, and the methods of using them as therapeutic agents for disease states in mammals characterized by abnormal thrombosis: 
Wherein:
R1 is selected from the group consisting of H, C1-6alkyl, C3-8cycloalkyl, C1-3alkylaryl, C1-3alkyl-C3-8cycloalkyl and aryl and R2 is H, or R1 and R2 are taken together to form a carbocyclic ring;
m is an integer from 0-2;
n is an integer from 0-6;
k is an integer from 0-1;
p is an integer from 0-1;
q is an integer from 0-3;
r is an integer from 0-4;
s is an integer from 0-1;
t is an integer from 0-4;
A is selected from the group consisting of R3, xe2x80x94NR3R4, 
where
R3, R4, R5 and R6 are independently selected from the group consisting of H, xe2x80x94OH, C1-6alkyl, aryl and C1-4alkylaryl; R1 is selected from the group consisting of H, C1-6alkyl, aryl and C1-4alkylaryl, or can be taken together with R5 or R6 to form a 5-6 membered ring; and R8 is selected from the group consisting of H, C1-6alkyl, aryl and C1-4alkylaryl, or can be taken together with R6 to form a 5-6 membered ring;
Q is selected from the group consisting of a direct link, xe2x80x94Sxe2x80x94, xe2x80x94SOxe2x80x94, xe2x80x94SO2xe2x80x94, xe2x80x94Oxe2x80x94, xe2x80x94SO2NHxe2x80x94, xe2x80x94NHSO2xe2x80x94, xe2x80x94COOxe2x80x94, xe2x80x94CONHxe2x80x94, C1-6alkyl, C3-8cycloalkyl, C2-6alkenyl, C2-6alkenylaryl, aryl and a five to ten membered heterocyclic ring system containing 1-4 heteroatoms selected from the group consisting of N, O and S;
D is selected from the group consisting of a direct link, xe2x80x94COxe2x80x94, xe2x80x94SO2xe2x80x94, xe2x80x94Oxe2x80x94COxe2x80x94, xe2x80x94NR9xe2x80x94SO2xe2x80x94 or xe2x80x94NR9xe2x80x94COxe2x80x94, where R9 is H, xe2x80x94OH, C1-6alkyl, aryl or C1-4alkylaryl;
E is selected from the group consisting of a direct link, C3-8cycloalkyl, aryl, or a five to ten membered heterocyclic ring system containing 1-4 heteroatoms selected from the group consisting of N, O and S;
G is selected from the group consisting of R10, xe2x80x94NR10OR11, 
where
R10, R11 , R12 and R13 are independently selected from the group consisting of H, xe2x80x94OH, C1-6alkyl, aryl and C1-4alkylaryl; R14 is selected from the group consisting of H, C1-6alkyl, aryl and C1-4alkylaryl, or can be taken together with R12 or R13 to form a 5-6 membered ring; and R15 is selected from the group consisting of H, C1-6alkyl, aryl and C1-4alkylaryl, or can be taken together with R13 to form a 5-6 membered ring; with the proviso that when G is R10, then E must contain at least one N atom;
K is selected from the group consisting of a direct link, C3-8cycloalkyl, C2-6alkenyl, C2-6alkenylaryl, aryl and a five to ten membered heterocyclic ring system containing 1-4 heteroatoms selected from the group consisting of N, O and S;
T is selected from the group consisting of R28, xe2x80x94NR28R29, 
where
R28, R29, R30 and R31 are independently selected from the group consisting of H, xe2x80x94OH, C1-6alkyl, aryl and C1-4alkylaryl; R32 is selected from the group consisting of H, C1-6alkyl, aryl and C1-4alkylaryl, or can be taken together with R30 or R31 to form a 5-6 membered ring; and R33 is selected from the group consisting of H, C1-6alkyl, aryl and C1-4alkylaryl, or can be taken together with R31 to form a 5-6 membered ring;
X and Y are independently selected from the group consisting of O and H2;
W is selected from the group consisting of H, 
where
R16 and R17 are independently selected from the group consisting of H, C1-3alkyl and aryl; and Z is selected from the group consisting of H, xe2x80x94COOR18, xe2x80x94CONR18R19, xe2x80x94CF3, xe2x80x94CF2CF3 and a group having the formula: 
where:
R18 and R19 are independently selected from the group consisting of H, C1-6alkyl, aryl and C1-4alkylaryl;
U is selected from the group consisting of xe2x80x94Oxe2x80x94, xe2x80x94Sxe2x80x94, xe2x80x94Nxe2x80x94 and xe2x80x94NHxe2x80x94; and
V is selected from the group consisting of xe2x80x94Oxe2x80x94, xe2x80x94Sxe2x80x94, xe2x80x94Nxe2x80x94 and xe2x80x94NHxe2x80x94; with the proviso that at least one of U or V is xe2x80x94Nxe2x80x94 or xe2x80x94NHxe2x80x94;
R20 is selected from the group consisting of H, C1-6alkyl, C2-6alkenyl, C0-6alkylaryl, C2-6alkenylaryl, C0-6alkylheterocyclo, C2-6alkenylheterocyclo, xe2x80x94CF3 and xe2x80x94CF2CF3;
J is selected from the group consisting of xe2x80x94Sxe2x80x94, xe2x80x94SOxe2x80x94, xe2x80x94SO2xe2x80x94,  Oxe2x80x94 and xe2x80x94NR21xe2x80x94, where R21 is selected from the group consisting of H, C1-6alkyl and benzyl; and
L is selected from the group consisting of: 
xe2x80x83a C6-10 heterocyclic ring system substituted by R24 and R25 and containing 1-4 heteroatoms selected from N, S and O;
where d is an integer from 0-2;
R22 and R23 are independently selected from the group consisting of H, C1-6alkyl, aryl, C1-6alkylaryl, xe2x80x94COOR26, xe2x80x94CONR26R27, xe2x80x94CN and xe2x80x94CF3;
R24 and R25 are independently selected from the group consisting of H, C1-6alkyl, aryl, C1-6alkylaryl, C1-4alkyloxy, halogen, xe2x80x94NO2, xe2x80x94NR26R27, xe2x80x94NR26COR27, xe2x80x94OR26, xe2x80x94OCOR26, xe2x80x94COOR26, xe2x80x94CONR26R27, xe2x80x94CN, xe2x80x94CF3, xe2x80x94SO2NR26R27 and C1-6alkyl-OR ; and
R26 and R27 are independently selected from the group consisting of H, C1-6alkyl, C1-3alkylaryl and aryl; and all pharmaceutically acceptable salts and optical isomers thereof.
A preferred embodiment of compounds of general structural formula I have the following stereochemistry: 
Preferred R1 substituents are H and C1-6alkyl; more preferably H and methyl; most preferably H. R2 is preferably H.
The integer xe2x80x9cmxe2x80x9d is preferably from 0-1; more preferably 0.
The integer xe2x80x9cnxe2x80x9d is preferably from 0-4.
The integer xe2x80x9cpxe2x80x9d is preferably 0.
The integer xe2x80x9cqxe2x80x9d is preferably from 1-2.
The integer xe2x80x9csxe2x80x9d is preferably 0.
The integer xe2x80x9ckxe2x80x9d is preferably 0.
The integer xe2x80x9crxe2x80x9d is preferably 3.
In the various xe2x80x9cAxe2x80x9d substituents, it is preferred that R3, R4, R5 and R6 are independently selected from the group consisting of H or C1-6alkyl; and are more preferably independently selected from the group consisting of H or methyl. It is also preferred that R7 is H, C1-6alkyl or taken together with R5 or R6 to form a 5-6 membered ring; and is more preferably H or methyl. It is also preferred that R8 is H, C1-6alkyl or taken together with R6 to form a 5-6 membered ring; and is more preferably H or methyl.
Preferred xe2x80x9cQxe2x80x9d substituents are a direct link, C1-4alkyl, C3-8cycloalkyl, aryl, or a five to ten membered heterocyclic ring system. More preferably, Q is C1-4alkyl, aryl, or a five to ten membered heterocyclic ring system.
D is preferably a direct link, xe2x80x94COxe2x80x94 or xe2x80x94SO2.
E is preferably a direct link.
In the xe2x80x9cGxe2x80x9d substituent, it is preferred that R10 R11, R12 and R13 are independently selected from the group consisting of H and C1-6alkyl, more preferably H and methyl.
K is preferably selected from the group consisting of a direct link, C3-8cycloalkyl, aryl and a five to ten membered heterocyclic ring system containing 1-4 heteroatoms selected from the group consisting of N, O and S;
In the various xe2x80x9cTxe2x80x9d substituents, it is preferred that R28, R29, R30 and R31 are independently selected from the group consisting of H or C1-6alkyl; and are more preferably independently selected from the group consisting of H or methyl. It is also preferred that R32 is H, C1-6alkyl or taken together with R30 or R31 to form a 5-6 membered ring; and is more preferably H or methyl. It is also preferred that R33 is H, C1-6alkyl or taken together with R30 to form a 5-6 membered ring; and is more preferably H or methyl.
X is preferably H2.
Y is preferably O.
W is preferably: 
where R16 is preferably H and R17 is preferably H.
Z is preferably H, xe2x80x94COOR18, xe2x80x94CONR18R19 or a group having the formula: 
For xe2x80x94COOR18, R18 is preferably H, C1-6alkyl or C1-4alkylaryl. For xe2x80x94CONR18R19, R18 is preferably H and R19 is preferably C1-4alkylaryl.
J is preferably xe2x80x94Sxe2x80x94, xe2x80x94Oxe2x80x94 or xe2x80x94NR21xe2x80x94, where R21 is preferably H or methyl, more preferably H.
L is preferably selected from the group consisting of: 
L is more preferably 
R24 and R25 are preferably independently selected from the group consisting of H, xe2x80x94Oxe2x80x94 R26, xe2x80x94COOR26, xe2x80x94CONR26R27 or xe2x80x94CF3; more preferably H.
When L is: 
then R22 is preferably H and R23 is preferably H.
When Z is: 
then R20 is preferably xe2x80x94CF3 or xe2x80x94CF2CF3.
In one preferred embodiment of the invention, k, p and s are 0, Y is O, R1 and R2 are H and W is xe2x80x94C(O)xe2x80x94Z. This is also illustrated as a preferred group of compounds defined by the general structural formula II as: 
A preferred embodiment of compounds of general structural formula II have the following stereochemistry: 
In another preferred embodiment of the invention, k, p, s and m are 0, Y is O, X is H2, R1 and R2 are H and W is xe2x80x94C(O)xe2x80x94Z. This is also illustrated as a preferred group of compounds defined by the general structural formula III as: 
A preferred embodiment of compounds of general structural formula III have the following stereochemistry: 
In another preferred embodiment of the invention, k, p, s and m are 0, r is 3, D is xe2x80x94SO2, Y is O, X is H2, R1 and R2 are H, E is a bond, G is: 
where R10, R12, R13 and R14 are all H, and W is xe2x80x94C(O)xe2x80x94Z. This is also illustrated as a preferred group of compounds defined by the general structural formula IV as: 
A preferred embodiment of compounds of general structural formula IV have the following stereochemistry: 
This invention also encompasses all pharmaceutically acceptable isomers, salts, hydrates and solvates of the compounds of formulas I, II, III and IV. In addition, the compounds of formulas I, II, III and IV can exist in various isomeric and tautomeric forms, and all such forms are meant to be included in the invention, along with pharmaceutically acceptable salts, hydrates and solvates of such isomers and tautomers.
The compounds of this invention may be isolated as the free acid or base or converted to salts of various inorganic and organic acids and bases. Such salts are within the scope of this invention. Non-toxic and physiologically compatible salts are particularly useful although other less desirable salts may have use in the processes of isolation and purification.
A number of methods are useful for the preparation of the salts described above and are known to those skilled in the art. For example, the free acid or free base form of a compound of one of the formulas above can be reacted with one or more molar equivalents of the desired acid or base in a solvent or solvent mixture in which the salt is insoluble, or in a solvent like water after which the solvent is removed by evaporation, distillation or freeze drying. Alternatively, the free acid or base form of the product may be passed over an ion exchange resin to form the desired salt or one salt form of the product may be converted to another using the same general process.
This invention also encompasses prodrug derivatives of the compounds contained herein. The term xe2x80x9cprodrugxe2x80x9d refers to a pharmacologically inactive derivative of a parent drug molecule that requires biotransformation, either spontaneous or enzymatic, within the organism to release the active drug. Prodrugs are variations or derivatives of the compounds of this invention which have groups cleavable under metabolic conditions. Prodrugs become the compounds of the invention which are pharmaceutically active in vivo, when they undergo solvolysis under physiological conditions or undergo enzymatic degradation. Prodrug compounds of this invention may be called single, double, triple etc., depending on the number of biotransformation steps required to release the active drug within the organism, and indicating the number of functionalities present in a precursor-type form. Prodrug forms often offer advantages of solubility, tissue compatibility, or delayed release in the mammalian organism (see, Bundgard, Design of Prodrugs, pp. 7-9, 21-24, Elsevier, Amsterdam 1985 and Silverman, The Organic Chemistry of Drug Design and Drug Action, pp. 352-401, Academic Press, San Diego, Calif., 1992). Prodrugs commonly known in the art include acid derivatives well known to practitioners of the art, such as, for example, esters prepared by reaction of the parent acids with a suitable alcohol, or amides prepared by reaction of the parent acid compound with an amine, or basic groups reacted to form an acylated base derivative. Moreover, the prodrug derivatives of this invention may be combined with other features herein taught to enhance bioavailability.
The following structures are illustrative of the compounds of the present invention and are not intended to be limiting in any manner. It is to be noted that in the compounds of the invention, certain substituents are present between two other substituents. For example, K is positioned between T and (CH2)t. Accordingly, substituents such as K are illustrated below as having two xe2x80x9cdanglingxe2x80x9d bonds, the bond on the left representing a direct link to substituent T and the bond on the right representing a direct link to (CH2)t. Therefore, the general formula of Txe2x80x94Kxe2x80x94(CH2)txe2x80x94 where K is phenyl can be written as: 
K, a phenyl group, would then be written as follows in the tables below: 
Other substituents in the table below may also be presented as having one or two similar xe2x80x9cdanglingxe2x80x9d bonds. It is understood that these represent direct links to the adjacent substituent(s). It is also understood that the compounds illustrated below can exist as other isomers, and the isomeric form illustrated herein is not intended to be limiting in any manner.
The invention encompasses compounds of general structural formula V, where R1 and R2 are H; m, p, k and s are 0; n, q and t are 1; r is 3; A and T are H; K and Q are phenyl;
D is xe2x80x94SO2xe2x80x94; E is a direct link; X is H2; Y is O; and G is 
where W is selected from the group consisting of: 
In addition, the invention encompasses compounds of general structural formula VI where R1 and R2 are H; m, p, k and s are 0; n and q are 1; r is 3; A and T are H; Q is phenyl; E is a direct link; D is xe2x80x94SO2xe2x80x94; G is xe2x80x94NHxe2x80x94C(NH)NH2; X is H2; Y is O; and W is 
In addition, the invention encompasses compounds of general structural formula VII where R1 and R2 are H; m, p, k and s are 0; n and q are 1; r is 3; A is H; Q is phenyl; W is 
E is a direct link; D is xe2x80x94SO2xe2x80x94; G is xe2x80x94NHxe2x80x94C(NH)NH2; X is H2 and Y is O:
In addition, the invention encompasses compounds of general structural formula VIII where R1 and R2 are H; m, p, k and s are 0; n and q are 1; r is 3; A is H; Q is phenyl; E is a direct link; D is xe2x80x94COxe2x80x94; G is xe2x80x94NHxe2x80x94C(NH)NH2; W is 
X is H2 and Y is O:
In addition, the invention encompasses compounds of general structural formula IX where R1 and R2 are H; m, p, k and s are 0; n and q are 1; r is 3; A is H; Q is phenyl; E and D are direct links; G is xe2x80x94NHxe2x80x94C(NH)NH2; W is 
X is H2 and Y is O:
In addition, the invention encompasses compounds of general structural formula X where R1 and R2 are H; p, k and s are 0; q and t are 1; r is 3; T is H; E is a direct link; D is xe2x80x94SO2xe2x80x94; K is phenyl; G is xe2x80x94NHxe2x80x94C(NH)NH2; W is 
X is H2 and Y is O:
In addition, the invention encompasses compounds of general structural formula XI where R1 and R2 are H; p, k, n and m are 0; q and t are 1; A and T are H; D is xe2x80x94SO2xe2x80x94; K and Q are phenyl; W is 
X is H2and Y is O:
In addition, the invention encompasses compounds of general structural formula XII where R1 and R2 are H; r is 3; s and m are 0; n and t are 1; E is a direct link; A and T are H; K and Q are phenyl; G is xe2x80x94NHxe2x80x94C(NH)NH2; and W is 
In addition, the invention encompasses compounds of general structural formula XIII where p, k, t and s are 0; q is 1; r is 3; T is H; E is a direct link; K and D are direct links; G is xe2x80x94NHxe2x80x94C(NH)NH2; W is 
X is H2 and Y is O:
A mentioned above, the compounds of this invention find utility as therapeutic agents for the states in mammals which have disorders of coagulation such as in the treatment or prevention of unstable angina, refractory angina, myocardial infarction, transient ischemic attacks, thrombotic stroke, embolic stroke, disseminated intravascular coagulation including the treatment of septic shock, deep venous thrombosis in the prevention of pulmonary embolism or the treatment of reocclusion or restenosis of reperfused coronary arteries. Further, these compounds are useful for the treatment or prophylaxis of those diseases which involve the production and/or action of factor Xa/prothrombinase complex. This includes a number of thrombotic and prothrombotic states in which the coagulation cascade is activated which include but are not limited to, deep venous thrombosis, pulmonary embolism, myocardial infarction, stroke, thromboembolic complications of surgery and peripheral arterial occlusion.
Accordingly, a method for preventing or treating a condition in a mammal characterized by undesired thrombosis comprises administering to the mammal a therapeutically effective amount of a compound of this invention. In addition to the disease states noted above, other diseases treatable or preventable by the administration of compounds of this invention include, without limitation, occlusive coronary thrombus formation resulting from either thrombolytic therapy or percutaneous transluminal coronary angioplasty, thrombus formation in the venous vasculature, disseminated intravascular coagulopathy, a condition wherein there is rapid consumption of coagulation factors and systemic coagulation which results in the formation of life-threatening thrombi occurring throughout the microvasculature leading to widespread organ failure, hemorrhagic stroke, renal dialysis, blood oxygenation, and cardiac catheterization.
The compounds of the invention also find utility in a method for inhibiting the coagulation biological samples, which comprises the administration of a compound of the invention.
The compounds of the present invention may also be used in combination with other therapeutic or diagnostic agents. In certain preferred embodiments, the compounds of this invention may be coadministered along with other compounds typically prescribed for these conditions according to generally accepted medical practice such as anticoagulant agents, thrombolytic agents, or other antithrombotics, including platelet aggregation inhibitors, tissue plasminogen activators, urokinase, prourokinase, streptokinase, heparin, aspirin, or warfarin. The compounds of the present invention may act in a synergistic fashion to prevent reocclusion following a successful thrombolytic therapy and/or reduce the time to reperfusion. These compounds may also allow for reduced doses of the thrombolytic agents to be used and therefore minimize potential hemorrhagic side-effects. The compounds of this invention can be utilized in vivo, ordinarily in mammals such as primates, (e.g. humans), sheep, horses, cattle, pigs, dogs, cats, rats and mice, or in vitro.
The biological properties of the compounds of the present invention can be readily characterized by methods that are well known in the art for example by the in vitro protease activity assays and in vivo studies to evaluate antithrombotic efficacy, and effects on hemostasis and hematological parameters, such as are illustrated in the examples.
Diagnostic applications of the compounds of this invention will typically utilize formulations in the form of solutions or suspensions. In the management of thrombotic disorders the compounds of this invention may be utilized in compositions such as tablets, capsules or elixirs for oral administration, suppositories, sterile solutions or suspensions or injectable administration, and the like, or incorporated into shaped articles. Subjects in need of treatment (typically mammalian) using the compounds of this invention can be administered dosages that will provide optimal efficacy. The dose and method of administration will vary from subject to subject and be dependent upon such factors as the type of mammal being treated, its sex, weight, diet, concurrent medication, overall clinical condition, the particular compounds employed, the specific use for which these compounds are employed, and other factors which those skilled in the medical arts will recognize.
Formulations of the compounds of this invention are prepared for storage or administration by mixing the compound having a desired degree of purity with physiologically acceptable carriers, excipients, stabilizers etc., and may be provided in sustained release or timed release formulations. Acceptable carriers or diluents for therapeutic use are well known in the pharmaceutical field, and are described, for example, in Remington""s Pharmaceutical Sciences, Mack Publishing Co., (A. R. Gennaro edit. 1985). Such materials are nontoxic to the recipients at the dosages and concentrations employed, and include buffers such as phosphate, citrate, acetate and other organic acid salts, antioxidants such as ascorbic acid, low molecular weight (less than about ten residues) peptides such as polyarginine, proteins. such as serum albumin, gelatin, or immunoglobulins, hydrophilic polymers such as polyvinylpyrrolidinone, amino acids such as glycine, glutamic acid, aspartic acid, or arginine, monosaccharides, disaccharides, and other carbohydrates including cellulose or its derivatives, glucose, mannose or dextrins, chelating agents such as EDTA, sugar alcohols such as mannitol or sorbitol, counterions such as sodium and/or nonionic surfactants such as Tween, Pluronics or polyethyleneglycol.
Dosage formulations of the compounds of this invention to be used for therapeutic administration must be sterile. Sterility is readily accomplished by filtration through sterile membranes such as 0.2 micron membranes, or by other conventional methods. Formulations typically will be stored in lyophilized form or as an aqueous solution. The pH of the preparations of this invention typically will be 3-11, more preferably 5-9 and most preferably 7-8. It will be understood that use of certain of the foregoing excipients, carriers, or stabilizers will result in the formation of cyclic polypeptide salts. While the preferred route of administration is by injection, other methods of administration are also anticipated such as orally, intravenously (bolus and/or infusion), subcutaneously, intramuscularly, colonically, rectally, nasally, transdermally or intraperitoneally, employing a variety of dosage forms such as suppositories, implanted pellets or small cylinders, aerosols, microencapsulation, oral dosage formulations and topical formulations such as ointments, drops and dermal patches. The compounds of this invention are desirably incorporated into shaped articles such as implants which may employ inert materials such as biodegradable polymers or synthetic silicones, for example, Silastic, silicone rubber or other polymers commercially available.
The compounds of the invention may also be administered in the form of liposome delivery systems, such as small unilamellar vesicles, large unilamellar vesicles and multilamellar vesicles. Liposomes can be formed from a variety of lipids, such as cholesterol, stearylamine or phosphatidylcholines.
The compounds of this invention may also be delivered by the use of antibodies, antibody fragments, growth factors, hormones, or other targeting moieties, to which the compound molecules are coupled. The compounds of this invention may also be coupled with suitable polymers as targetable drug carriers. Such polymers can include polyvinylpyrrolidinone, pyran copolymer, polyhydroxy-propyl-methacrylamide-phenol, polyhydroxyethyl-aspartamide-phenol, or polyethyleneoxide-polylysine substituted with palmitoyl residues. Furthermore, compounds of the invention may be coupled to a class of biodegradable polymers useful in achieving controlled release of a drug, for example polylactic acid, polyglycolic acid, copolymers of polylactic and polyglycolic acid, polyepsilon caprolactone, polyhydroxy butyric acid, polyorthoesters, polyacetals, polydihydropyrans, polycyanoacrylates and cross linked or amphipathic block copolymers of hydrogels. Polymers and semipermeable polymer matrices may be formed into shaped articles, such as valves, stents, tubing, prostheses and the like.
Therapeutic compound liquid formulations generally are placed into a container having a sterile access port, for example, an intravenous solution bag or vial having a stopper pierceable by hypodermic injection needle.
Therapeutically effective dosages may be determined by either in vitro or in vivo methods. For each particular compound of the present invention, individual determinations may be made to determine the optimal dosage required. The range of therapeutically effective dosages will be influenced by the route of administration, the therapeutic objectives and the condition of the patient. For injection by hypodermic needle, it may be assumed the dosage is delivered into the body""s fluids. For other routes of administration, the absorption efficiency must be individually determined for each compound by methods well known in pharmacology. Accordingly, it may be necessary for the therapist to titer the dosage and modify the route of administration as required to obtain the optimal therapeutic effect. The determination of effective dosage levels, that is, the dosage levels necessary to achieve the desired result, will be readily determined by one skilled in the art. Typically, applications of compound are commenced at lower dosage levels, with dosage levels being increased until the desired effect is achieved.
The compounds of the invention can be administered orally or parenterally in an effective amount within the dosage range of about 0.1 to 100 mg/kg, preferably about 0.5 to 50 mg/kg and more preferably about 1 to 20 mg/kg on a regimen in a single or 2 to 4 divided daily doses and/or continuous infusion.
Typically, about 5 to 500 mg of a compound or mixture of compounds of this invention, as the free acid or base form or as a pharmaceutically acceptable salt, is compounded with a physiologically acceptable vehicle, carrier, excipient, binder, preservative, stabilizer, dye, flavor etc., as called for by accepted pharmaceutical practice. The amount of active ingredient in these compositions is such that a suitable dosage in the range indicated is obtained.
Typical adjuvants which may be incorporated into tablets, capsules and the like are binders such as acacia, corn starch or gelatin, and excipients such as microcrystalline cellulose, disintegrating agents like corn starch or alginic acid, lubricants such as magnesium stearate, sweetening agents such as sucrose or lactose, or flavoring agents. When a dosage form is a capsule, in addition to the above materials it may also contain liquid carriers such as water, saline, or a fatty oil. Other materials of various types may be used as coatings or as modifiers of the physical form of the dosage unit. Sterile compositions for injection can be formulated according to conventional pharmaceutical practice. For example, dissolution or suspension of the active compound in a vehicle such as an oil or a synthetic fatty vehicle like ethyl oleate, or into a liposome may be desired. Buffers, preservatives, antioxidants and the like can be incorporated according to accepted pharmaceutical practice.
The compounds of the present invention may be synthesized by either solid or liquid phase methods described and referenced in standard textbooks, or by a combination of both methods. These methods are well known in the art. See, Bodanszky, xe2x80x9cThe Principles of Peptide Synthesisxe2x80x9d, Hafner, et al., Eds., Springer-Verlag, Berlin, 1984.
Starting materials used in any of these methods are commercially available from chemical vendors such as Aldrich, Sigma, Nova Biochemicals, Bachem Biosciences, and the like, or may be readily synthesized by known procedures.
Reactions are carried out in standard laboratory glassware and reaction vessels under reaction conditions of standard temperature and pressure, except where otherwise indicated. The reaction products are isolated and purified by conventional methods, typically by solvent extraction into a compatible solvent. The products may be further purified by column chromatography such as reversed-phase HPLC, or other appropriate methods. Characterization of the compounds can easily be accomplished by ion-spray mass spectrometry.
During the synthesis of these compounds, the functional groups of the amino acid derivatives used in these methods are protected by blocking groups to prevent side reactions during the coupling procedure. Examples of suitable blocking groups and their use are described in xe2x80x9cThe Peptides: Analysis, Synthesis, Biologyxe2x80x9d, Academic Press, Vol. 3 (Gross, et al., Eds., 1981) and Vol. 9 (1987), the disclosures of which are incorporated herein by reference.
The compounds of the invention may be preferably prepared by
a) coupling the carboxylic acid of formula (a) to the amine of formula (b) 
xe2x80x83by the standard amide bond formation strategies, or
b) where X is H2 D is xe2x80x94SO2xe2x80x94 or xe2x80x94COxe2x80x94, reacting the cyclic amine of formula (c) with the sulfonyl halide of formula (d) or with the carboxylic acid of formula of (e) through standard amide bond or sulfonamide formation strategies, 
The compounds of formula (b) wherein W is H can be prepared by the methods disclosed in WO 96/01338; WO 96/24609; Feng, et al., WO 96/31504; and WO 96/32110, the disclosures of which are incorporated herein by reference.
The compounds of formula (b) where W is a boron containing compound can be prepared by the methods disclosed in J. Org. Chem. 60:3717-3722 (1995) and de Nanteuil, et al., EP 688,788, the disclosures of which are incorporated herein by reference.
The compounds of formula (b) where W is xe2x80x94C(O)xe2x80x94Z, where Z is H may be prepared by the methods disclosed in WO 93/15756, supra; Vlasuk, et al., WO 94/17817; Abelman, et al., WO 94/21673; Webb, et al., WO 94/08941; Veber, et al., WO 94/25051; Levy, et al., WO 95/35312; Semple, et al., WO 95/35313; Abelman, et al., WO 95/28420; and Abelman, et al., WO 96/19493, the disclosures of which are incorporated herein by reference.
The compounds of formula (b) where W is xe2x80x94C(O)xe2x80x94Z, where Z is xe2x80x94COOR18 or xe2x80x94COCONR18R19 may be prepared by the methods disclosed in WO 94/25051, supra, WO 94/08941, supra, and WO 94/21673, supra, the disclosures of which are incorporated herein by reference.
The compounds of formula (b) where W is xe2x80x94C(O)xe2x80x94Z, where Z is xe2x80x94CF3 or xe2x80x94CF2CF3 may be prepared by the methods disclosed in Schacht, et al., GB 2287027, the disclosure of which is incorporated herein by reference.
The compounds of formula (b) where W is xe2x80x94C(O)xe2x80x94Z, where Z is: 
and J is O, xe2x80x94SOxe2x80x94 or xe2x80x94SO2xe2x80x94 can be readily synthesized by the methods disclosed in Costanzo, et al., U.S. Pat. No. 5,523,308; Di Maio, et al., WO 96/19483; U.S. Pat. No. 5,164,371; J. Am. Chem. Soc. 114: 1854-1863 (1992); J. Med. Chem. 38:76-85 (1995); and J. Med. Chem. 37:3492-3502 (1994). Lastly, fragments where J is xe2x80x94NR21xe2x80x94, where R21 is H, C1-6alkyl or benzyl, can be synthesized by techniques illustrated in J. Med. Chem. 37:3492-3502 (1994). All of these references are incorporated herein by reference.
The compounds of formula (b) wherein W is xe2x80x94C(O)xe2x80x94Z, where Z is: 
and U and V are the various substituents (xe2x80x94Oxe2x80x94, xe2x80x94Sxe2x80x94, xe2x80x94Nxe2x80x94, xe2x80x94NHxe2x80x94) may be prepared by the methods disclosed in J. Med. Chem. 38: 1355-1371 (1995) and J. Med. Chem. 37: 2421-2436 (1994), the disclosures of which are incorporated herein by reference.
The starting compounds of formula (a), (c), (d) and (e) are either known compounds or can be produced by known methods (Heitsch, et al., Canadian Patent No. 2,071,744; Sugihara, et al., Canadian Patent No. 2,126,026; Baker, et al., EP 365,992; U.S. Pat. No. 4,251,438; Carr, et al., U.S. Pat. No. 4,341,698; Goldman, et al., U.S. Pat. No. 5,120,718; Biswanath, et al., U.S. Pat. No. 5,164,388; Duggan, et al., U.S. Pat. No. 5,281,585; Sugihara, et al., U.S. Pat. No. 5,294,713; Bovy, et al., WO 95/06038; WO 95/35308; J. Chem. Soc. Perkin Trans. I 1687-1689 (1989); and Int. J. Peptide Protein Res. 37:468-475 (1991)) or prepared by the methods shown in the following reaction formulae.
The following reaction schemes are more specific illustrations of the above reaction formulae, where xe2x80x9cPxe2x80x9d is a protecting group for the amino group and xe2x80x9cRxe2x80x9d is a protecting group for the carboxylic acid such as Me, Et, Bn or t-Bu. The chemical reactions described in each scheme can easily be modified and combined with other techniques that are well known in the art to produce other compounds within the scope of the invention. 